In gene therapy, genes are introduced into an organism for the purpose of expressing them in the organism. The regulation of the expression of these genes is important to the prophylactic or therapeutic effect of gene therapy.
Patent Applications PCT/GB95/02000, PCT/EP95/03370, PCT/EP95/03371, PCT/EP95/03368, PCT/EP95/03339 describe gene expression regulators. These regulators comprise an activator sequence whose function is, for example, the cell-specific or virus-specific activation of basal transcription. The DNA sequence of this activator sequence is linked, by its 3' end, to the 5' end of a promoter module. The structural gene is in turn linked, by its 5' end, to the 3' end of the promoter module.
The promoter module comprises nucleic acid sequences which bind to the transcription factors of the CDF and CHF or E2F and CHF families. In the G0 and G1 phases of the cell cycle, this binding leads to inhibition of the upstream activator sequence and consequently to inhibition of the transcription of the structural gene which is located downstream (i.e., in the direction of transcription).
In the G0 and G1 phases of cell division, the DNA contained in the cell is in the diploid state. The cell is at rest in the G0 phase and is inhibited in its cell-cycle progression in the G1 phase. The G1 phase is followed by the S phase, in which DNA synthesis takes place and in which the genome is replicated. There then follows the G2 phase, in which the cell is in the tetraploid state. The G2 phase is followed by cell division (mitosis or "M phase"). The daughter cells pass into the G0 or G1 states, and so on.
Consequently, combining a cell-specific or virus-specific activator sequence with a promoter module which inhibits this activator sequence in the G0 and G1 phases makes it possible to regulate the expression of a structural gene both in a cell-specific or virus-specific manner and in a cell-cycle-specific manner (i.e., restricted to the S and G2 phases).
The combination of an activator sequence and a promoter module is termed a "chimeric" promoter. While there are numerous possibilities for applying chimeric promoters in gene therapy, the shortcomings of these promoters also impose a number of limitations.
Examples of these limitations include:
a weak activator sequence, which leads to insufficient transcription of the structural gene, PA1 the use of an activator sequence which cannot be adequately inhibited in a cell-cycle-dependent manner by the chosen promoter module, PA1 the restriction to two (for example cell or virus-specific and cell-cycle-specific) regulators of the transcription of the structural gene, and PA1 inadequate retention of the transcription product in the cell nucleus PA1 and/or inadequate intracellular transport of the transcription product of the structural gene which has been introduced into the cell.